Immunoreactive distribution of gonadotropin-inhibitory hormone precursor, RFRP, in a temperate bat species (Eptesicus fuscus).

Gonadotropin-inhibitory hormone (GnIH, also known RFRP-3 in mammals) is an important regulator of the hypothalamic-pituitary-gonadal axis and downstream reproductive physiology. Substantial species differences exist in the localization of cell bodies producing RFRP-3 and patterns of fiber immunoreactivity in the brain, raising the question of functional differences. Many temperate bat species exhibit unusual annual reproductive patterns. Male bats upregulate spermatogenesis in late spring which is asynchronous with periods of mating in the fall, while females have the physiological capacity to delay their reproductive investment over winter via sperm storage or delayed ovulation/fertilization. Neuroendocrine mechanisms regulating reproductive timing in male and female bats are not well-studied. We provide the first description of RFRP-precursor peptide of GnIH -expression and localization in the brain of any bat using a widespread temperate species (Eptesicus fuscus, big brown bat) as a model. RFRP mRNA expression was detected in the hypothalamus, testes, and ovaries of big brown bats. Cellular RFRP-immunoreactivity was observed within the periventricular nuclei, dorsomedial nucleus of the hypothalamus, arcuate nucleus (Arc), and median eminence (ME). As in other vertebrates, RFRP fiber immunoreactivity was widespread, with the greatest density observed in the hypothalamus, preoptic area, Arc, ME, midbrain, and thalamic nuclei. Putative interactions between RFRP-ir fibers and gonadotropin-releasing hormone (GnRH) cell bodies were observed in 16% of GnRH-immunoreactive cells, suggesting direct regulation of GnRH via RFRP signaling. This characterization of RFRP distribution contributes to a deeper understanding of bat neuroendocrinology, which serves as foundation for manipulative approaches examining changes in reproductive neuropeptide signaling in response to environmental and physiological challenges within, and among, bat species.

Clinical utility of urinary gonadotrophins in hypergonadotrophic states as Turner syndrome.

Background Girls with Turner syndrome (TS) are at an increased risk of primary ovarian insufficiency (POI). Good correlation between serum and urinary gonadotrophins exists in children assessed for disorders of puberty, but there is little evidence of their reliability in hypergonadotropic states. Objectives To determine whether there was a correlation between serum and urinary Luteinising Hormone (uLH) and Follicle-Stimulating Hormone (uFSH) in hypergonadotrophic states, and whether uFSH could suggest an ovarian failure in TS as Anti-Mullerian Hormone (AMH). Patients and Methods Retrospective cohort study of 37 TS girls attending the paediatric TS clinic in Glasgow between February 2015 and January 2019, in whom 96 non-timed spot urine samples were available with a median age at time of sample of 12.89 years (3.07-20.2 years). uLH and uFSH were measured by chemiluminescent microparticle immunoassay. Simultaneous serum gonadotrophins and AMH were available in 30 and 26 girls, respectively. AMH <4 pmol/L was considered indicative of ovarian failure. Results A strong correlation was found between serum LH and uLH (r 0.860, P<0.001) and serum FSH and uFSH (r 0.905, p<0.001). Among patients≥10 years not on oestrogen replacement, ROC curve identified uFSH as a reasonable marker for AMH<4 pmol/L uFSH of >10.85 U/L indicates an AMH <4 pmol/L with 75% sensitivity and 100 % specificity (AUC 0.875)with similar ability as serum FSH (AUC 0.906). Conclusion uLH and uFSH are non-invasive, useful and reliable markers of ovarian activity in hypergonadotropic states as TS. uFSH could provide an alternative to AMH (in centres which are limited by availability or cost) in revealing ovarian failure and requirement for oestrogen replacement in pubertal induction.

Hormonal, metabolic and inflammatory circulating biomarker profiles in obese and non-obese Brazilian middle-aged women.

AIM: To investigate circulating hormonal, metabolic and inflammatory biomarker profiles in obese and non-obese middle-aged women. METHODS: A total of 110 women, aged 40-60 years, were included in this cross-sectional study. Patients were allocated, according to the occurrence of menopause and body mass index (BMI), into four groups: PM0 (premenopausal non-obese), PM1 (premenopausal obese), M0 (postmenopausal non-obese), and M1 (postmenopausal obese). Serum levels of gonadotropins, sex hormones, lipid markers, leptin, hs-CRP and interleukin-6 were obtained using either colorimetric or immunoenzymatic assays. Univariate and correlation analyses were performed among all clinical and laboratorial parameters. Principal component analysis was used to characterize subsets of biomarkers, which had their discriminatory capacity tested using discriminant function analysis. RESULTS: Levels of gonadotropins and female sex hormones were similar between PM0 and PM1 and between M0 and M1 (p > 0.05), all of them varied between PM0 and M0 (p < 0.05), but only estradiol was significantly altered in the comparison between PM1 and M1 (p = 0.027). Regarding metabolic markers, leptin was lower in PM0 than in M0 (p = 0.010) and higher in M1 than in M0 (p = 0.046). In premenopausal women, BMI correlated only to leptin, while it correlated to several other markers in postmenopausal women. A combination of FSH and leptin serum levels significantly discriminated the four groups (Wilks's lambda < 0.001, in canonical functions 1 and 2). CONCLUSION: A combined analysis of hormonal biomarkers may potentially distinguish obese from non-obese women with distinct menopause status. Further research is thus required to clarify the clinical significance of such findings.

Gonadotropins and Their Analogs: Current and Potential Clinical Applications.

The gonadotropin receptors LH receptor and FSH receptor play a central role in governing reproductive competency/fertility. Gonadotropin hormone analogs have been used clinically for decades in assisted reproductive therapies and in the treatment of various infertility disorders. Though these treatments are effective, the clinical protocols demand multiple injections, and the hormone preparations can lack uniformity and stability. The past two decades have seen a drive to develop chimeric and modified peptide analogs with more desirable pharmacokinetic profiles, with some displaying clinical efficacy, such as corifollitropin alfa, which is now in clinical use. More recently, low-molecular-weight, orally active molecules with activity at gonadotropin receptors have been developed. Some have excellent characteristics in animals and in human studies but have not reached the market-largely as a result of acquisitions by large pharma. Nonetheless, such molecules have the potential to mitigate risks currently associated with gonadotropin-based fertility treatments, such as ovarian hyperstimulation syndrome and the demands of injection-based therapies. There is also scope for novel use beyond the current remit of gonadotropin analogs in fertility treatments, including application as novel contraceptives; in the treatment of polycystic ovary syndrome; in the restoration of function to inactivating mutations of gonadotropin receptors; in the treatment of ovarian and prostate cancers; and in the prevention of bone loss and weight gain in postmenopausal women. Here we review the properties and clinical application of current gonadotropin preparations and their analogs, as well as the development of novel orally active, small-molecule nonpeptide analogs.

Enzyme-linked immunosorbent assay of relaxin-like gonad-stimulating peptide in the starfish Patiria (Asterina) pectinifera.

A relaxin-like gonad-stimulating peptide (RGP) from starfish Patiria (Asterina) pectinifera is the first identified invertebrate gonadotropin for final gamete maturation. Recently, we succeeded in obtaining specific antibodies against P. pectinifera RGP (PpeRGP). In this study, the antibodies were used for the development of a specific and sensitive enzyme-linked immunosorbent assay (ELISA) for the measurement of PpeRGP. A biotin-conjugated peptide that binds to peroxidase-conjugated streptavidin is specifically detectable using 3,3',5,5'-tetramethylbenzidine (TMB)/hydrogen peroxide as a substrate; therefore, biotin-conjugated RGP (biotin-PpeRGP) was synthesized chemically. Similarly to PpeRGP, synthetic biotin-PpeRGP bound to the antibody against PpeRGP. In binding experiments with biotin-PpeRGP using wells coated with the antibody, a displacement curve was obtained using serial concentrations of PpeRGP. The ELISA system showed that PpeRGP could be measured in the range 0.01-10pmol per 50µl assay buffer. On the contrary, the B-chains of PpeRGP, Asterias amurensis RGP, Aphelasterias japonica RGP, and human relaxin showed minimal cross-reactivity in the ELISA, except that the A-chain of PpeRGP affected it slightly. These results strongly suggest that this ELISA system is highly specific and sensitive with respect to PpeRGP.

Cambios hormonales después del tratamiento de cáncer de próstata localizado. Comparación entre radioterapia de haz externo y prostatectomía radical / Hormonal changes after localized prostate cancer treatment. Comparison between external beam radiation therapy and radical prostatectomy

Objetivos: Determinar la influencia de la prostatectomía radical (PR) y de la radioterapia externa (RT) sobre el eje hipotálamo-hipofisario de 120 pacientes con cáncer de próstata clínicamente localizado tratados con PR o RT exclusiva. Material y métodos: Estudiamos 120 pacientes con cáncer de próstata localizado. Noventa y dos pacientes recibieron PR y 28 RT exclusiva. Medimos los niveles séricos de hormona luteinizante, hormona folículo estimulante (FSH), testosterona total (T), testosterona libre y estradiol basalmente y a los 3 y 12 meses tras completar el tratamiento. Resultados: Los pacientes sometidos a PR eran más jóvenes y presentaban mayor volumen prostático (64,3 vs. 71,1 años, p < 0,0001 y 55,1 vs. 36,5 g, p < 0,0001; respectivamente). No encontramos diferencias en los niveles hormonales basales. Los niveles de hormona luteinizante y FSH eran significativamente superiores en los pacientes tratados con RT a los 3 meses (hormona luteinizante 8,54 vs. 4,76 U/l, FSH 22,96 vs. 8,18 U/l, p < 0,0001) y los niveles de T y testosterona libre significativamente inferiores (T 360,3 vs. 414,83 ng/dl, p 0,039; FT 5,94 vs. 7,5 pg/ml, p 0,018). A los 12 meses los niveles de FSH permanecían significativamente superiores en los pacientes tratados con RT comparado con pacientes tratados con PR (21,01 vs. 8,51 U/l, p < 0,001) y los niveles de T permanecían significativamente inferiores (339,89 vs. 402,39 ng/dl, p 0,03). Conclusiones: El tratamiento del cáncer de próstata influye en el eje hipotálamo-hipofisario. La influencia parece más importante en los pacientes tratados con RT. Necesitamos más estudios que eluciden el papel que la próstata puede jugar como órgano endocrino

Objective: To determine the influence of radical prostatectomy (RP) and external beam radiation therapy (EBRT) on the hypothalamic pituitary axis of 120 men with clinically localized prostate cancer treated with RP or EBRT exclusively. Materials and methods: 120 patients with localized prostate cancer were enrolled. Ninety two patients underwent RP and 28 patients EBRT exclusively. We measured serum levels of luteinizing hormone, follicle stimulating hormone (FSH), total testosterone (T), free testosterone, and estradiol at baseline and at 3 and 12 months after treatment completion. Results: Patients undergoing RP were younger and presented a higher prostate volume (64.3 vs. 71.1 years, p < 0.0001 and 55.1 vs. 36.5 g, p < 0.0001; respectively). No differences regarding serum hormonal levels were found at baseline. Luteinizing hormone and FSH levels were significantly higher in those patients treated with EBRT at three months (luteinizing hormone 8,54 vs. 4,76 U/l, FSH 22,96 vs. 8,18 U/l, p < 0,0001) while T and free testosterone levels were significantly lower (T 360,3 vs. 414,83 ng/dl, p 0,039; free testosterone 5,94 vs. 7,5 pg/ml, p 0,018). At 12 months FSH levels remained significantly higher in patients treated with EBRT compared to patients treated with RP (21,01 vs. 8,51 U/l, p < 0,001) while T levels remained significantly lower (339,89 vs. 402,39 ng/dl, p 0,03). Conclusions: Prostate cancer treatment influences the hypothalamic pituitary axis. This influence seems to be more important when patients with prostate cancer are treated with EBRT rather than RP. More studies are needed to elucidate the role that prostate may play as an endocrine organ

Development of a homologous radioimmunoassay for red seabream follicle stimulating hormone and regulation of gonadotropins by GnRH in red seabream, Pagrus major.

Using a recombinant chimeric single-chain follicle stimulating hormone (FSH), we established a radioimmunoassay (RIA) for red seabream (Pagrus major) FSH (pmFSH) which became a powerful tool for studying reproductive physiology. We studied the profiles in plasma and pituitary concentrations of FSH and luteinizing hormone (LH) during sexual maturation. A pre-established RIA for red seabream LH was used for the LH measurements. The regulation of FSH and LH secretion from the pituitary was investigated using a gonadotropin-releasing hormone analog (GnRHa) in vivo and in vitro. Marked differences in plasma and pituitary FSH levels were observed between males and females; pituitary FSH content in males was much higher than that in females during all seasons, and plasma FSH levels in males were high during the spawning season, whereas those in females were unchanged. In contrast, plasma and pituitary levels of LH were elevated before and during the spawning season in males and females. Injecting or implanting (cholesterol pellet) a GnRHa into adult and juvenile red seabream resulted in significant increases in plasma LH concentrations; however, no significant change was observed in plasma FSH. Moreover, GnRHa stimulated only LH secretion in an in vitro experiment using dispersed pituitary cells. The discrete FSH and LH secretion profiles revealed suggest differential roles for the two gonadotropins during red seabream gametogenesis. In addition, the marked difference in pituitary FSH levels in males and females suggests the relative significance of FSH in male reproduction.

Symptoms of gonadal dysfunction are more predictive of hypopituitarism than nonspecific symptoms in screening for pituitary dysfunction following moderate or severe traumatic brain injury.

OBJECTIVE: The economic and logistic burden of screening for hypopituitarism following moderate/severe traumatic brain injury (TBI) is considerable. A key recommendation in published guidelines is to prioritize for screening those patients with symptoms suggestive of pituitary dysfunction. The purpose of this study was to evaluate the utility of targeted screening for hypopituitarism in long-term survivors after moderate/severe TBI using referrals on the basis of symptoms. DESIGN: In group 1 (G1), consecutive, unselected patients were screened from the Irish National Neurosurgery Centre, whereas in group 2 (G2) patients were targeted based on the presence of symptoms suggestive of pituitary dysfunction. PATIENTS: A total of 137 patients (113 male) were systematically screened (G1) and compared to 112 patients (77 male) referred for pituitary evaluation on the basis of suggestive symptoms (G2). MAIN OUTCOME MEASURES: The rate of GH, ACTH, gonadotrophin (GT), TSH and ADH deficiency was compared among groups. RESULTS: Patients referred with menstrual dysfunction had more GH (50% vs 11%, P = 0·001), ACTH (60% vs 14%, P < 0·0001), GT (90% vs 16%, P < 0·0001) deficiency and any pituitary hormone deficit (80% vs 33%, P = 0·003) than G1. Men with symptoms of hypogonadism had more GH (33% vs 11%, P = 0·003), GT (58% vs 16%, P < 0·0001) and TSH (16% vs 1%, P = 0·03) deficiency than G1. Patients with nonspecific symptoms were no more likely to have hypopituitarism than those consecutively screened. CONCLUSIONS: Symptoms of hypogonadism are sufficiently predictive of hypopituitarism to justify screening for hypopituitarism after moderate/severe TBI. Nonspecific symptoms of hypopituitarism are no more predictive than unselected screening.

The effect of water quality on the immunoreactivity of stress-response cells and gonadotropin-secreting cells in the pituitary gland of Nile tilapia, Oreochromis niloticus.

The present experiments investigated the effect of water quality characteristics on the condition factor, the ovarian activity, cortisol level, and the immunoreactivity of stress-response cells (adrenocorticotropic hormone; ACTH- and melanin stimulating hormone; MSH- and somatolactin; SL- secreting cells) and gonadotropin (GTH)-secreting cells in the pituitary gland of Nile tilapia, Oreochromis niloticus. After 3 months of exposure to mixtures of water from different sources (Tap and Lake Manzalah waters), with high levels of minerals and heavy metals, water quality affected the number, size, and immunostaining of stress-response-immunoreactive (ir) cells and GTH-ir cells, which showed a dramatic decrease in their size. The integrated optical density (IOD) of immunoreactivity of MSH- and GTH- cells was significantly increased; however, it was significantly decreased for ACTH- and SL- cells. Also, high levels of cortisol were observed in females exposed to waters with high concentrations of minerals and heavy metals. In parallel, low values of gonadosomatic index (GSI%) and the ovarian histology revealed a decrease of maturing follicles concomitant with an increase of atretic follicles in females exposed to Lake Manzalah polluted water. Taken together, the increased activity of stress-response-ir pituitary cells, serum cortisol level and ovarian atretic follicles in response to elevated concentrations of minerals and heavy metals, supports the possible role of ACTH, MSH, and SL in the adaptive stress response of fish. Therefore, minerals and heavy metals must be considered when discussing tilapia aquaculture status.

Current trends in mass spectrometry of peptides and proteins: Application to veterinary and sports-doping control.

Detection of misuse of peptides and proteins as growth promoters is a major issue for sport and food regulatory agencies. The limitations of current analytical detection strategies for this class of compounds, in combination with their efficacy in growth-promoting effects, make peptide and protein drugs highly susceptible to abuse by either athletes or farmers who seek for products to illicitly enhance muscle growth. Mass spectrometry (MS) for qualitative analysis of peptides and proteins is well-established, particularly due to tremendous efforts in the proteomics community. Similarly, due to advancements in targeted proteomic strategies and the rapid growth of protein-based biopharmaceuticals, MS for quantitative analysis of peptides and proteins is becoming more widely accepted. These continuous advances in MS instrumentation and MS-based methodologies offer enormous opportunities for detection and confirmation of peptides and proteins. Therefore, MS seems to be the method of choice to improve the qualitative and quantitative analysis of peptide and proteins with growth-promoting properties. This review aims to address the opportunities of MS for peptide and protein analysis in veterinary control and sports-doping control with a particular focus on detection of illicit growth promotion. An overview of potential peptide and protein targets, including their amino acid sequence characteristics and current MS-based detection strategies is, therefore, provided. Furthermore, improvements of current and new detection strategies with state-of-the-art MS instrumentation are discussed for qualitative and quantitative approaches.

Genetics of male infertility.

Genetics play an important role in the evaluation of the infertile male. The current limitations of classifying the genetic contribution to male infertility and the importance of phenotyping men are discussed, and the core concepts necessary to interpret most genetic studies are reviewed. The current genetic assays used clinically are discussed in detail. The use and interpretation of the cystic fibrosis transmembrane receptor assay are examined in the context of men with clinical bilateral absence of the vas deferens, a karyotype and Klinefelter syndrome, and Y chromosome microdeletions. The role of hormones and epigenetics in evaluating the genetic reproductive potential of men is discussed briefly. A summary of what the field might look like in 2034 is presented.

Diagnóstico clínico y bioquímico del déficit androgénico en el varón adulto / Clinical and biochemical diagnosis of androgenic deficit in adult men

El diagnóstico de déficit de testosterona (DT) en el varón adulto se basa en la constatación de una serie de síntomas o signos clínicos indiciarios asociada a unos niveles bajos de T sérica verificados. El DT puede resultar de múltiples causas: fallo primario del testiculo, déficit de su estimulación por LH (secundario) o de ambos fenómenos conjuntamente (combinado o mixto). La expresión clínica del DT depende de cuando se inicie. Si es antes de completarse el desarrollo puberal resulta en un fenotipo eunucoide relativamente fácil de detectar. Cuando aparece después de la pubertad su expresión clínica es proteiforme; sus síntomas son inespecíficos, confundiéndose con los del envejecimiento y diversos trastornos crónicos. Con más frecuencia se afecta la función sexual, reflejándose en falta de deseo y dificultades de erección. Los cuestionarios propuestos para su detección muestran poca especificidad y débil correlación con la T plasmática, desaconsejándose como instrumentos diagnósticos aunque pueden ayudar para cuantificar los síntomas y monitorizar la respuesta a un ulterior tratamiento. La exploración física suele aportar pocos datos significativos, pero puede encontrar signos de un DT de inicio temprano no detectado antes. Es preceptiva la medición del nivel plasmático total de testosterona total (TT). En varones adultos el límite inferior del intervalo normal de referencia se sitúa en torno a 10 nmol/L, aunque varía entre laboratorios. Los niveles de TT en el intervalo de ± 20% alrededor de este límite representan una “zona gris” que debe ser aclarada. Para ello se suelen calcular sus fracciones libre (TL) o biodisponible (TBD) a partir de los valores de TT, SHBG y albúmina (AU)

Confirmado el SDT, la búsqueda de su etiología con pruebas adicionales que incluyen la medición de gonadotropinas y prolactina ayuda a orientar la intervención terapéutica. Se resumen aquí en un algoritmo diagnóstico. Se necesitan métodos para evaluar fiablemente la sensibilidad androgénica de cada individuo (AU)

The diagnosis of testosterone deficit (TD) in adult men is made upon the finding of consistent symptoms or signs associated with confirmed low Testosterone levels. TD can be caused by primary testicular failure, decreased LH secretion (secondary) or a combination of both (mixed type). The clinical features of TD depend on the age of onset. Before completion of pubertal development it results in eunuchoid traits that can be easily detected. The clinical expression of adult onset TD is protean, with varied symptoms and signs often overlapping with those of aging and several chronic conditions. Sexual problems as low desire and difficult erections are frequently reported by TD patients. Several symptom questionnaires have been proposed for detecting TD. However, poor specificity and weak correlation with serum testosterone make them unsuitable as diagnostic tools, although their scores can help for monitoring future treatments. Physical examination usually yields few significant data but it can help for detecting unidentified cases of early onset TD. Measuring serum total testosterone(TT) levels is mandatory. It is available in most laboratories although commonly used methods are not fully satisfactory. In adult men the lower limit for the normal reference range is around 10 nmol/ L, but this figure can vary among laboratories. “Grey zone” values falling within a ± 20% interval around this limit need further clarification. To this aim most hospital laboratories can calculate the free (FT) or bioavailable fractions (BioT) of TT, SHBG, and albumin levels. When TD diagnosis is confirmed, a number of additional tests, such as measuring gonadotropins and prolactin, should be used to find out its etiology. They are summarized here in a diagnostic algorithm. There is a need for laboratory tests giving a reliable estimate of the androgen sensitivity of each subject (AU)

Gonadotropin and sex steroid levels in HIV-infected premenopausal women and their association with subclinical atherosclerosis in HIV-infected and -uninfected women in the women's interagency HIV study (WIHS).

BACKGROUND: HIV-infected women may experience prolonged amenorrhea, suggesting altered gonadotropin and sex hormone levels. However, the impact of these endocrine disruptions on atherosclerosis has not been evaluated in women living with, or at risk for, HIV infection. We investigated the association of sex hormone and gonadotropin concentrations with subclinical atherosclerosis in HIV-infected and -uninfected premenopausal women in the Women's Interagency HIV Study. METHODS: Using B-mode ultrasound, the common carotid artery intima-media thickness and distensibility were measured once. Cycle-specific FSH, total estradiol (E2), and inhibin-B concentrations were measured in 584 (414 HIV infected, 170 HIV uninfected) women. Random concentrations of total T, dehydroepiandrosterone sulphate, and SHBG were measured in 1094 (771 HIV infected, 323 HIV uninfected) women. The endocrine analytes were measured at or before the ultrasound visit. Sex hormones, FSH, and SHBG concentrations were compared between HIV-infected and -uninfected women using nonparametric testing. Linear regression models were used to evaluate the association of sex hormones, FSH, and SHBG with carotid artery intima-media thickness and distensibility adjusted for confounders. Separate analyses were conducted by HIV status. RESULTS: Compared with HIV-uninfected women, E2, T, and dehydroepiandrosterone sulphate concentrations were significantly lower and SHBG was higher in HIV-infected women. Adjusted for the confounders, T was significantly positively associated with distensibility (ß-estimate = .04, P = .0005) among HIV-infected women, and the magnitude of association did not differ by CD4 cell count. E2 was significantly positively associated with distensibility among HIV-infected women with CD4 count less than 350 cells/µL. CONCLUSIONS: HIV-infected women had reduced estrogen and androgen compared with HIV-uninfected premenopausal women. T deficiency is linked with carotid artery stiffness, regardless of immune suppression, whereas E2 deficiency is linked with carotid stiffness among immunocompromised HIV-infected premenopausal women. Further research is warranted to understand the impact of endocrine dysregulation on the accelerated cardiovascular disease risk in HIV-infected women.

Fabrication of stable antibody-modified field effect transistors using electrical activation of Schiff base cross-linkages for tumor marker detection.

In this paper, we present a method of fabricating a rigid antibody-immobilized surface using electric activation of a glutaraldehyde (GA)-modified aminopropylsilyl surface for stable antibody-modified field effect transistors (FETs). Electric activation of the GA-modified gate surface of the FET reduces Schiff bases, which are easily hydrolyzed and collapsed, formed between GA and 3-aminopropyltriethoxysilane, resulting in preventing the immobilized antibodies from desorbing from the surface. The lack of Raman peaks that could be assigned to a Schiff base after the electrical activation of the GA-modified surface indicated that the electric activation had reduced the Schiff base. The use of the antibody-modified FETs has three advantages for the detection of antigens: increased sensitivity, distinct recognition ability, and improved reproducibility. A tumor marker, alpha-fetoprotein (AFP), was quantitatively detected up to a concentration of 10 ng/mL using the antibody-modified FET. The detection ability of the FET accomplished a cutoff value of hepatic cancer. The quantitative detection of AFP in a solution with contaminating proteins was also demonstrated. This electric activation method is applicable to other antibody-modified FETs.

The relationship between plasma levels of the endocannabinoid, anandamide, sex steroids, and gonadotrophins during the menstrual cycle.

OBJECTIVE: To further investigate the relationship between plasma anandamide (AEA), sex steroids, and gonadotrophins to improve our understanding of how AEA may be involved in human fertility. DESIGN: Cross-sectional and longitudinal study. SETTING: University Hospital of Leicester NHS Trust, Leicester Royal Infirmary. PATIENT(S): Healthy premenopausal and postmenopausal volunteers. INTERVENTION(S): UPLC-MS/MS-measured plasma AEA and ELISA-measured serum FSH, LH, estradiol, and progesterone levels at five different phases of the menstrual cycle and postmenopause. MAIN OUTCOME MEASURE(S): Plasma AEA, serum steroids and gonadotrophins. RESULT(S): Changes in AEA levels were similar in the two cohorts. The mean +/- SEM levels in the early follicular phase (0.89 +/- 0.06) for the cross-sectional cohort and the longitudinal cohort (0.73 +/- 0.03) were higher than those in the late follicular phase (0.77 +/- 0.09 cross-sectional; 0.63 +/- 0.08 longitudinal). The highest AEA levels were measured at ovulation (1.38 +/- 0.14 and 1.33 +/- 0.16) and the lowest level was measured in the late luteal phase (0.66 +/- 0.07 and 0.56 +/- 0.06). There was a statistically significant positive correlation between AEA, estradiol (P=0.0015), LH (P<0.0001) and FSH levels but not progesterone (P=0.022). CONCLUSION(S): Peak plasma AEA occurred at ovulation and positively correlated with estradiol and gonadotrophin levels suggesting that these may be involved in the regulation of AEA levels.

Molecular characterization of Chinese sturgeon gonadotropins and cellular distribution in pituitaries of mature and immature individuals.

Chinese sturgeon (Acipenser sinensis) is a rare and endangered species, and also an important resource for the sturgeon aquaculture industry. To understand molecular characterization of Chinese sturgeon gonadotropins (GTHs), we cloned the full-length cDNAs of gonadotropin subunits common alpha (GTH-alpha), follicle-stimulating hormone (FSH) and luteinizing hormone (LH) from a pituitary cDNA library of mature female. Two subtypes of GTH-alpha were identified. The nucleotide sequences of A. sinensis common alpha I (AsGTH-alpha I), common alpha II (AsGTH-alpha II), FSHbeta (AsFSHbeta) and LHbeta (AsLHbeta) subunit cDNAs are 345, 363, 387 and 414bp in length, and encode mature peptides of 115, 121, 129 and 138aa, respectively. Then, three polyclonal antibodies were prepared from the in vitro expressed AsGTH-alpha I, AsFSHbeta and AsLHbeta mature proteins, respectively. Significant expression differences were revealed between immature and mature sturgeon pituitaries. Western blot detection and immunofluoresence localization revealed the existence of three-gonadotropin subunits (AsGTH-alpha, AsFSHbeta and AsLHbeta) in mature sturgeon pituitaries, but only AsFSHbeta was detected in immature individual pituitaries during early stages in the sturgeon life, and obvious difference was observed between males and females. In males, AsFSHbeta was expressed in 4-year-old individuals, whereas in females, AsFSHbeta was just expressed in 5-year-old individuals.

Comparative gene expression of gonadotropins (FSH and LH) and peptide levels of gonadotropin-releasing hormones (GnRHs) in the pituitary of wild and cultured Senegalese sole (Solea senegalensis) broodstocks.

The Senegalese sole (Solea senegalensis) is a valuable flatfish for aquaculture, but it presents important reproductive problems in captivity. Spawning is achieved by wild-caught breeders but cultured broodstocks fail to spawn spontaneously and, when they do, eggs are unfertilized. To gain knowledge on the physiological basis underlying this reproductive dysfunction, this study aimed at analyzing comparative hormone levels between wild and cultured broodstocks at the spawning season. The Senegalese sole gonadotropin (GTH) subunits, FSHbeta, LHbeta and GPalpha, were cloned and qualitative (in situ hybridization) and quantitative (real-time PCR) assays developed to analyze pituitary GTH gene expression. In females, FSHbeta and GPalpha mRNA levels were higher in wild than in cultured broodstocks, whereas in males all three subunits were highest in cultured. By ELISA, three GnRH forms were detected in the pituitary, displaying a relative abundance of GnRH2>GnRH1>GnRH3. All GnRHs were slightly more abundant in wild than cultured females, whereas no differences were observed in males. Plasma levels of vitellogenin and sex steroids were also analyzed. Results showed endocrine differences between wild and cultured broodstocks at the spawning period, which could be related to the endocrine failure of the reproductive axis in cultured breeders.

Reproductive alterations in male rats exposedperinatally to Solanum lycocarpum fruits / Alterações reprodutivas em ratos machos expostos perinatalmente aos frutos da Solanum lycocarpum

This work employed pregnant rats treated with Solanum lycocarpum unripe fruits (10% in diet) from gestation day (GD) 06 to post-natalday (PND) 07, for the evaluation of the sperm number, daily sperm production and epididymal sperm transit time of the male offspring at PND 60 and PND 90. No differences were observed in the daily sperm production (DSP) and sperm number in the testis of the exposed males at PND 60 and PND 90. Also, no alterations were observed in sperm transit time in the caput epididymis of the exposed males at PND 60 and PND 90. However, a reduced sperm transit time was observed in the corpus/cauda epididymis of the experimental males at PND 90. The last data may explain the reduced sperm number observed in the corpus/cauda epididymis of the experimental male rats at PND 90. These data show that the male rats exposed to S.lycocarpum fruits during gestation did not present alterations in testis sperm production and number, however the sperm transit time through epididymis was impaired, resulting in a decreased number of spermatozoa in epididymis cauda. We conclude that S. lycocarpummay cause imbalance on hypothalamus-pituitary gland axis

Ratas prenhes foram tratadas do dia 06 da gestação (GD 06) ao dia 07 pós-natal (PND 07) com frutos verdes secos e moídos da Solanum lycocarpum (10% na ração). Após nascimento das ninhadas, foi avaliado na prole masculina adulta aos 60 e 90 dias de vida, o número de espermátides e a produção espermática diária nos testículos e o tempo de trânsito espermático no epidídimo. A exposição não foi capaz de promover alterações na produção espermática diária (DSP) e no número de espermátides produzidas pelo testículo dos ratos expostos aos frutos verdes da S. lycocarpum durante a gestação e início da lactação. Não foram observadas alterações no tempo de trânsito espermático na cabeça do epidídimo, porém, foi constatado menor número de espermatozóides no corpo/cauda do epidídimo nos machos experimentais com 90 dias de vida, provavelmente resultante do menor tempo de trânsito espermático observado no corpo/cauda do epidídimo aos PND 90. Estes dados sugerem que a exposição de ratos aos frutos verdes da S. lycocarpum durante a gestação e início dalactação, não foi suficiente para promover alterações na produção mas sim no trânsito espermático, indicando possível alteração no eixo hormônio liberador das gonadotrofinas hipotálamo-hipófise-gônada

Gonadotropins and female sex steroid hormones in cyst fluid and serum from patients with ovarian tumors.

The objective of the present study was to determine the concentrations of LH, FSH, 17beta-estradiol and progesterone in ovarian cyst fluid and serum from patients with benign and malignant ovarian tumors and to assess the correlation of the gonadotropin and female sex steroid hormone concentrations with menopausal and tumor status. Ovarian cyst fluid and blood samples were prospectively collected from 103 patients with ovarian tumors. Seventy-four of the patients had benign ovarian tumors while 29 patients had malignant ovarian tumors. Malignant ovarian tumors showed significantly higher LH and FSH cyst fluid concentrations compared to concentrations from patients with benign tumors. Within the malignant subset, LH and FSH concentrations correlated with increasing FIGO stage and grade. Furthermore, LH and FSH cyst fluid concentrations showed strong correlations (r > 0.62) with serum concentrations in case of malignant tumors, especially in postmenopausal women, but not in case of benign tumors. The highest gonadotropin concentrations were observed in cyst fluid from malignant ovarian tumors. The most probable explanation for this is an increased vascular permeability within the cysts. Supportive evidence for such an increased vascular permeability is our previous finding of significantly higher VEGF concentrations in cyst fluid from malignant ovarian tumors. The possibility of ectopic production of LH and FSH by malignant ovarian tissue cannot completely be ruled out.

Menarquia extraordinariamente precoz como signo inicial de una pubertad precoz central idiopática en una niña de 15 meses / Extraordinarily early menarche as an initial sign of central idiopathic precocious puberty in a 15 month old child

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